A fluorescence optical system illuminates a fluorophore-labeled target with light whose wavelength content falls within the absorption band and collects light whose wavelength content is in the emission band. An emission filter placed in front of a detector filters light that is not in the emission band. One challenge with emission filters is that unwanted photon rejection depends on the angle at which light traverses the filter. Specifically, as the angle of incidence increases, the transmission/reflection of the filter shifts to lower wavelengths. Accordingly, even if the field of view is a single point that provides an axial ray at a 0 degree angle, other rays of the same light beam will pass through the filter at non-0 degree angles and, accordingly, may experience different amounts of filtering.
This situation is addressed in Hwang et al., “The influence of improved interference filter performance for molecular imaging using frequency domain photon migration measurements,” Optical Tomography and Spectroscopy of Tissue VI, SPIE vol. 5693, pp. 503-512. Hwang et al. describes an optical system in which a collimator is placed between imaging optics and an emission filter. The collimator ensures that all rays in a light beam originating from a certain point in the image field will pass through the filter at a 0 degree angle and, thus, will receive the same type of filtering. However, if a relatively large field of view is used, light beams emanating from the edge of the field, while still collimated, will pass through the filter at an angle. This results in different amounts of excitation leakage across the field.
There is a need, therefore, for a fluorescence filtering method and system that will overcome this problem.